Highly efficient production of transfructosylating enzymes using low-cost sugarcane molasses by A. pullulans FRR 5284.

Fructooligosaccharides (FOS) are a type of important prebiotics and produced by transfructosylating enzymes. In this study, sugarcane molasses was used as the substrate for production of transfructosylating enzymes by Aureobasidium pullulans FRR 5284. NaNO3 was a superior nitrogen source to yeast extract for production of transfructosylating enzymes by A. pullulans FRR 5284 and decreasing the ratio of NaNO3 to yeast extract nitrogen from 1:0 to 1:1 resulted in the reduction of the total transfructosylating activity from 109.8 U/mL to 82.5 U/mL. The addition of only 4.4 g/L NaNO3 into molasses-based medium containing 100 g/L mono- and di-saccharides resulted in total transfructosylating activity of 123.8 U/mL. Scale-up of the A. pullulans FRR 5284 transfructosylating enzyme production process from shake flasks to 1 L bioreactors improved the enzyme activity and productivity to 171.7 U/mL and 3.58 U/mL/h, 39% and 108% higher than those achieved from shake flasks, respectively. Sucrose (500 g/L) was used as a substrate for extracellular, intracellular, and total A. pullulans FRR 5284 transfructosylating enzymes, with a maximum yield of 61%. Intracellular, extracellular, and total A. pullulans FRR 5284 transfructosylating enzymes from different production systems resulted in different FOS profiles, indicating that FOS profiles can be controlled by adjusting intracellular and extracellular enzyme ratios and, hence prebiotic activity.

Transformation of sugarcane molasses into fructooligosaccharides with enhanced prebiotic activity using whole-cell biocatalysts from Aureobasidium pullulans FRR 5284 and an invertase-deficient Saccharomyces cerevisiae 1403-7A.

Fructooligosaccharides (FOS) can be used as feed prebiotics, but are limited by high production costs. In this study, low-cost sugarcane molasses was used to produce whole-cell biocatalysts containing transfructosylating enzymes by Aureobasidium pullulans FRR 5284, followed by FOS production from molasses using the whole-cells of A. pullulans. A. pullulans in molasses-based medium produced cells and broth with a total transfructosylating activity of 123.6 U/mL compared to 61.0 and 85.8 U/mL in synthetic molasses-based and sucrose-based media, respectively. It was found that inclusion of glucose in sucrose medium reduced both transfructosylating and hydrolytic activities of the produced cells and broth. With the use of pure glucose medium, cells and broth had very low levels of transfructosylating activities and hydrolytic activities were not detected. These results indicated that A. pullulans FRR 5284 produced both constitutive and inducible enzymes in sucrose-rich media, such as molasses while it only produced constitutive enzymes in the glucose media. Furthermore, treatment of FOS solutions generated from sucrose-rich solutions using an invertase-deficient Saccharomyces yeast converted glucose to ethanol and acetic acid and improved FOS content in total sugars by 20-30%. Treated FOS derived from molasses improved the in vitro growth of nine probiotic strains by 9-63% compared to a commercial FOS in 12 h incubation. This study demonstrated the potential of using molasses to produce FOS for feed application.